Fast screening immunoassay of sulfonamides in commercial fish samples

Abstract

An indirect competitive enzyme-linked immunosorbent assay (ELISA) was developed in plate to detect three sulfonamide residues (sulfamerazine (SMR), sulfadimetoxine (SDM), and sulfadiazine (SDZ)) in gilthead sea bream (Sparus aurata) samples. Different extraction methodologies--using methanol/water 1:1 (v/v) + ethylene diamine tetraacetic acid (EDTA) 0.5% (m/v), acetonitrile, phosphate-buffered saline (PBS) 10 mmol L(-1) pH 7 and acetate buffer 100 mmol L(-1) pH 5--and cleanup steps, based on solid-phase extraction (C(18), SCX, Si) or liquid extraction with hexane, were assayed. As optimum, a fast and simple method using acetonitrile was selected to extract the sulfonamide residues from the edible muscle of fish. Due to matrix effects, a standard addition calibration curve in fish extract is necessary for quantification purposes. Sulfonamide-free samples were spiked at different concentration levels (between 30 and 90 ng g(-1), 5-15 ng mL(-1) in plate) and average recoveries (n = 8), ranging from 71% to 95%, 65% to 79%, and 72% to 95%, were obtained for SMR, SDM, and SDZ, respectively. The assay detection limits for these antibiotics were lower than 100 microg kg(-1) (maximum residue level established by the European Union). The accuracy was evaluated by spiking blank fish extracts at different concentrations (10-40 ng mL(-1), 5-20 ng mL(-1) in plate), and the relative errors ranged between +/-20%. Finally, in order to confirm the utility of the developed ELISA as a screening methodology, fish samples from different supermarkets were analyzed, and results were compared with those obtained by a validated high-performance liquid chromatography (HPLC) method. The correlation between the results obtained by both ELISA and HPLC methods is satisfactory.

Establishment of a monoclonal antibody-based indirect enzyme-linked immunosorbent assay for the detection of trimethoprim residues in milk, honey, and fish samples

“…Organic solvent in the supernatant was evaporated under a nitrogen stream. The remainder of samples were re-suspended in 4 mL of PBS-T containing 10% acetonitrile and filtered through 0.22 µm membrane (Cháfer-Pericás et al, 2010). This procedure diluted the fish tissue 4× to final TMP concentrations of 2, 4, and 8 µg kg −1 .…”

“…Fish tissue (5 g) in a 50-mL centrifuge tube was spiked with TMP resulting in three different TMP concentrations (8, 16, and 32 µg kg −1 ). The spiked fish tissues were placed in an ultrasonic bath for 5 min to thoroughly disperse TMP into the matrix (Cháfer-Pericás, Maquieira, Puchades, Miralles, & Moreno, 2010). Twenty-five milliliter of acetonitrile was added into the sample and mixed on a votex for 10 min.…”

Automated Determination of Sulfadiazine in Water, Fish Plasma and Muscle by HPLC with On‐Line Column‐Switching

“…[23,24]. There are several previous reports issued on rapid screening of drugs in serum and muscle after deproteinisation and extraction of the samples [22,[25][26][27][28]. In a recent review, the applicability of column-switching HPLC systems for trace analysis of organic compounds was described by Fern andez-Ramos et al [29].…”

Rapid automated method for on-site determination of sulfadiazine in fish farming: a stainless steel veterinary syringe coated with a selective membrane of PVC serving as a potentiometric detector in a flow-injection-analysis system

“…Sensors relying on immunoassays [10,11] are quite effective but are quite expensive for routine and continuous monitoring. Chemical sensors relying on charged and neutral carriers are also effective and have been proposed [12,13].…”